CD4 T cells are crucial to the control of Mycobacterium tuberculosis infection. This major T cell subset is traditionally thought of as providing help for B cells and producing cytokines for activating the effector cells of the immune system. In tuberculosis, the CD4 T cells produce IFN-3, a key cytokine to controlling the infection. However, other cells produce IFN-3 and our previous work demonstrated that depletion of CD4 T cells during chronic M. tuberculosis infection in a mouse did not substantially reduce the IFN-3 production in the lungs. Thus, we hypothesized that CD4 T cells likely have roles in addition to the production of IFN-3 for controlling M. tuberculosis infection. In this renewal proposal, we will identify key effector functions of CD4 T cells that must be stimulated for a robust immune response against this pathogen. This includes a close examination of the importance of IFN-3 throughout infection, and particularly the role of IFN-3 produced by CD4 T cells (Aim 1). In addition, we will explore other effector mechanisms of CD4 T cells, including additional cytokine production, cytotoxic functions, and effects on CD8 T cells (Aim 2). The CD4 memory response against M. tuberculosis is complex. Although most people infected with the organism do not develop disease, in large part due to ongoing immune responses, exogenous reinfection does occur. In our animal models, protection (vaccine or infection- induced) equals only approximately a 10-fold reduction in bacterial numbers. Thus, the immune responses that develop during primary infection or following vaccination may be flawed, or the ability of the challenge infection to appropriately recall the T cells to the lungs may be impaired. We will explore these important concepts in Aim 3. In the past funding period, we have developed and obtained exciting and novel reagents that will enhance our ability to accomplish the stated goals. These include mouse strains, M. tuberculosis strains, and new immunologic assays. This project is the result of a long standing collaboration between two labs: JoAnne Flynn's lab at the University of Pittsburgh and John Chan's lab at Albert Einstein. We work closely on all aspects of the project, perform experiments in both of our labs, publish together, share data and reagents. PUBLIC HEALTH RELEVANCE Tuberculosis, caused by the microbe Mycobacterium tuberculosis, kills 2 million people every year, even though much of the world's population is vaccinated at birth with BCG. We do not understand how to induce effective immunity against tuberculosis. A type of immune cell, the CD4 T cell, is clearly important in control of tuberculosis, since AIDS patients, who have reduced CD4 T cells, are extremely susceptible to this infection. This application will address how CD4 T cells contribute to immunity to tuberculosis, and provide insight into improving the CD4 T cell response, and possibly improving vaccines against this disease.